Method and system for real-time fluorescent determination of trace elements

ABSTRACT

A system for real-time fluorescent determination of trace elements comprising transport means for moving a plurality of samples, means for generating a plurality of incident radiation pulses of different wavelength, means for illuminating at least a respective one of the samples with at least a respective one of the radiation pulses during the movement of the samples, means for detecting the resultant fluorescence emitted from each of the samples and control means for controlling the movement of the samples and the incident radiation.

This application is filed pursuant to 35 U.S.C. § 371 as a United StatesNational Phase Application of International Application No.PCT/US01/26892, filed Aug. 29, 2001, which claims priority from UnitedStates Provisional Application No. 60/228,673, filed Aug. 29, 2000.

FIELD OF THE PRESENT INVENTION

The present invention relates generally to spectroscopy systems. Moreparticularly, the invention relates to a method and system for real-timefluorescent determination of trace elements.

BACKGROUND OF THE INVENTION

Beginning in the early 1970's, it was found that certain medicines couldbe administered in dry-powder form directly to the lungs by inhalationthrough the mouth or inspiration through the nose. This process allowsthe medicine to bypass the digestive system, and in some instances,allows smaller doses to be used to achieve the same desired results asorally ingested medicines.

Various metered dose powdered inhalers (“MDPI”) or nebulizers thatprovide inhalable mists of medicines are known in the art. Illustrativeis the devices disclosed in U.S. Pat. Nos. 3,507,277; 4,147,166 and5,577,497.

Most of the prior art MDPI devices employ powdered medicine contained ina gelatin capsule. The capsules are typically pierced and a metered doseof the powdered medicine is slowing withdrawn by partial vacuum, forcedinspiration of the user or by centrifugal force.

Several MDPI devices, such as that disclosed in U.S. Pat. No. 5,873,360employs a foil blister strip. Referring to FIG. 1, the foil blisterstrip 10 includes a plurality of individual, sealed blisters (orpockets) 12 that encase the powdered medicine. The blisters 12 aresimilarly pierced during operation to release the metered dose ofpowdered medicine.

As will be appreciated by one having ordinary skill in the art, theprovision of an accurate dosage of medicine in each capsule or blisteris imperative. Indeed, the U.S. Government mandates 100% inspection ofMDPI formulations to ensure that the formulations contain the properamount of prescribed medicine or drug(s).

Various technologies have been employed to analyze MDPI formulations(i.e., pharmaceutical compositions), such as X-ray diffraction,high-pressure liquid chromatography (HPLC) and UV/visible analysis.There are, however, numerous drawbacks associated with the conventionaltechnologies.

A major drawback of the noted technologies is that most require samplesto be collected from remote, inaccessible, or hazardous environments,and/or require extensive sampling that is time consuming andprohibitively costly. A further drawback is that detection of minuteamounts of trace elements, including the active ingredient or drug(s),is often difficult or not possible.

It is therefore an object of the present invention to provide a methodand system for high-speed, real-time, on-line fluorescent assessment ofactive ingredients and trace elements.

It is another object of the present invention to provide a method andsystem for high-speed, real-time, on-line fluorescent detection ofminute amounts of active ingredients and trace elements.

It is yet another object of the present invention to provide a methodand system for high-speed, real-time, on-line fluorescent determinationof the identity and concentration of active ingredients and traceelements.

SUMMARY OF THE INVENTION

In accordance with the above objects and those that will be mentionedand will become apparent below, the system for real-time fluorescentdetermination in accordance with this invention comprises means formoving a plurality of samples along a sample path; means for generatinga plurality of incident radiation pulses of different wavelength; meansfor illuminating at least a respective one of the samples with at leasta respective one of the radiation pulses during the movement of thesamples, the radiation pulse having a suitable range of fluorescenceradiation wavelengths; means for detecting the resultant fluorescenceemitted from each of the samples; and first control means incommunication with the moving means and the incident radiationgenerating means for synchronizing the means for illuminating each ofthe samples with the moving means.

The method for real-time fluorescent determination in accordance withthis invention generally comprises moving a plurality of said sampleshaving at least one element along a sample path; generating a pluralityof incident radiation pulses of different wavelength; illuminating atleast a respective one of the samples with at least a respective one ofthe radiation pulses during movement of the samples, the radiation pulsehaving a suitable range of fluorescence radiation wavelengths; detectingthe resultant fluorescence emitted from each of said samples; andcomparing the detected resultant fluorescence characteristics withstored fluorescence characteristics of predetermined elements and/oractive ingredients to identify the element or elements in the samples.

BRIEF DESCRIPTION OF THE DRAWINGS

Further features and advantages will become apparent from the followingand more particular description of the preferred embodiments of theinvention, as illustrated in the accompanying drawings, and in whichlike referenced characters generally refer to the same parts or elementsthroughout the views, and in which:

FIG. 1 is a perspective view of a prior art foil blister strip;

FIG. 2 is a side plan view of the foil blister strip shown in FIG. 1;

FIG. 3 is a flow chart of a conventional blister strip manufacturingprocess;

FIG. 4 is a schematic illustration of the fluorescence detection meansaccording to the invention;

FIG. 5 is a partial plan view of the radiation transmission means,illustrating the travel of the incident and emitted radiation accordingto the invention;

FIG. 6 is a further flow chart of a conventional foil blister stripmanufacturing process, illustrating the incorporation of thefluorescence detection means according to the invention;

FIG. 7 is a perspective view of a conventional conveyor and thefluorescence detection means according to the invention;

FIG. 8 is a partial section, front plan view of the conveyor andfluorescence detection means shown in FIG. 7; and

FIGS. 9 and 10 are graphs of incident radiation versus emissionradiation for prepared compounds, illustrating the detection of lowconcentration active trace elements according to the invention.

DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENT

The method and system of the present invention substantially reduces oreliminates the drawbacks and shortcomings associated with prior artmethods and systems for in-situ detection and analysis of traceelements. As discussed in detail below, the system generally includesfluorescence detection means adapted to provide high-speed, accurate,in-situ determination of the presence, identity and concentration oftrace elements and, in particular, active ingredients in pharmaceuticalcompositions. By the term “trace element”, it is meant to mean andinclude an ingredient, component or element of a pharmaceuticalcomposition or MDPI formulation having a relative concentration (i.e., %of total) of less than 0.5%, including, but not limited to, an activeingredient or element and medicament.

Referring first to FIG. 4, there is shown a schematic illustration ofthe fluorescence detection means (designated generally 20) of theinvention. The fluorescence detection means 20 generally comprises atleast one radiation transmission means 22 adapted to provide incidentradiation to the sample 14 and detect the fluorescence (emission)radiation from the sample 14, and first control means 24. As illustratedin FIG. 3, the first control means 24 preferably includes a light source26 for providing the desired wavelength of light or radiation to theradiation transmission means 22 via line 23 a, an analyzer 28 foranalyzing the emission radiation detected by the radiation transmissionmeans 22, which is communicated to the analyzer 28 via line 23 b, andstorage means for storing fluorescence characteristics of known elements(or ingredients) for subsequent comparison with detected emission(fluorescence) radiation from the sample(s) 14.

As discussed in detail below, the fluorescence detection means 20further includes second control means 29 preferably in communicationwith the light source 26, analyzer 28 and conveyor system 50 forsynchronizing the movement of the samples 14 on the conveyor system 50with the incident radiation transmission and detection of the resultantemission radiation (See FIG. 7).

As is well known in the art, for fluorescence measurements, it isnecessary to separate the emission (or emitted) radiation from theincident radiation. This is typically achieved by measuring the emissionradiation at right angles to the incident radiation.

However, as illustrated in FIG. 5, in a preferred embodiment of thepresent invention, the emission radiation, I_(o), is measured (ordetected) along a line I″ that is substantially coincident to the lineI′ defined by the travel of the incident radiation I. According to theinvention, the wavelength of the emission radiation I_(o) is “redshifted” to an upper frequency.

It is further well established that the relationship between the traceelement concentration and the fluorescence intensity (i.e., emissionradiation) can be derived from Beer's Law, i.e.,F=ΦP _(o)(1−10^(−∝bc))  EQ-1where:

-   -   F=Fluorescence Intensity    -   P₀=Power of incident radiation    -   ∝=Molar Absorbtivity    -   b=Path length    -   c=Sample concentration (moles/liter)    -   Φ=Quantum yield—a proportionality constant and a measure of the        fraction of absorbed photons that are converted into fluorescent        photons.

It is thus evident that the quantum yield, Φ, is generally less than orequal to unity. It is further evident from Eq. 1 that if the product ∝bcis large, the term 10^(−∝bc) becomes negligible compared to 1, and Fbecomes constant:F=ΦP_(o)  Eq. 2

Conversely, if the product ∝bc is small (≦0.01), it can be shown (i.e.,Taylor expansion series) that the following provides a goodapproximation of the fluorescence intensity:F=2.303ΦP_(o)∝bc  Eq. 3

Accordingly, for low concentrations of trace elements, the fluorescenceintensity is directly proportional to the concentration. Thefluorescence intensity is also directly proportional to the incidentradiation.

Since the noted relationships hold for concentrations up to a few partsfor million, Eq. 3 is preferably employed in the method of the inventionto determine the concentration of the trace element(s) detected by thefluorescence detection means 22.

Referring now to FIG. 3, there is shown a flow chart of a conventionalblister strip process, illustrating the primary steps involved in themanufacture of a foil blister strip. According to the process, the basefoil is fed from a coil 30 to the forming operation 32.

After the blisters 12 are formed on the strip 10 (see FIGS. 1 and 2),the strip 10 is inspected for defects 34 and, in particular, pin holes.Each blister 12 on the strip 10 is then filled 38 with a desired MDPIformulation or pharmaceutical composition.

After filling, the strip 10 is subjected to a second inspection 40. Thesecond inspection typically comprises a complete chemical analysis ofthe pharmaceutical composition to determine the presence of allingredients or elements and the respective concentrations thereof

As discussed above, the noted inspection 40 typically involves theremoval of a sample, transfer of the sample to an off-line location orfacility, and HPLC or UV/vis analysis. The operation is thus timeconsuming and expensive.

After the inspection 40, the appropriate code is applied 42 to the strip12. The strip is then transferred to a storage roll.

Referring now to FIG. 6, there is shown a further flow chart of theabove discussed blister strip process, illustrating the incorporation ofthe fluorescence detection means 20 of the invention. As illustrated inFIG. 6, the fluorescence detection means 20 is preferably disposedbetween the filling 38 and sealing 40 operations.

As will be appreciated by one having ordinary skill in the art, thefluorescence detection means 20 of the invention is readily adaptable tomost processes. Further, due to the inherent accuracy and tightspecifications (that are possible by virtue of the detection means 20),the conventional inspection (i.e., analysis) operation/step 38 can beeliminated. However, as illustrated in FIG. 6, the fluorescencedetection means 20 can also be employed in conjunction with theconventional inspection operation 38 (shown in phantom).

Referring to FIGS. 7 and 8, the fluorescence detection means 20 of theinvention will now be described in detail. Referring first to FIG. 7,there is shown a conventional conveyor system 50 adapted to facilitatethe transfer of two blister strips 10 a, 10 b to the above notedoperations 30, 32, 36, 20, 40, 42. As illustrated in FIG. 7, theradiation transmission means 22 is disposed proximate the conveyorsystem 50 and, hence, blister strips 10 a, 10 b positioned thereon.

In a preferred embodiment of the invention, the radiation transmissionmeans 22 comprises a J. Y. Horiba fluorometer that is adapted to providetwo lines of incident radiation (or incident radiation pulses) 25 a, 25b. According to the invention, the first line of incident radiation 25 ais directed toward and substantially perpendicular to the first blisterstrip 10 a and, hence, sample path (designated generally SP₁) and thesecond line of incident radiation 25 b is directed toward andsubstantially perpendicular to the second sample path (designatedgenerally SP₂ ). In additional envisioned embodiments of the invention,not shown, the radiation transmission means 22 is adapted to provide oneline of incident radiation (e.g., 25 a) to facilitate a single (ratherthan dual) blister strip process.

In a preferred embodiment of the invention, the first control means 24generates and provides a plurality of incident radiation pulses ofdifferent wavelengths, preferably in the range of 200 to 800 nm.According to the invention, at least a respective one of the samples 14is illuminated with at least a respective one of the incident radiationpulses as it traverses a respective sample path SP₁, SP₂. In a preferredembodiment, each sample 14 passing under the radiation transmissionmeans 22 is illuminated with incident radiation over a pre-determined,suitable range of wavelengths capable of inducing a fluorescenceresponse in at least one target element (or ingredient).

Applicants have found that the noted incident radiation wavelength rangewill induce a definitive fluorescence response in trace elements and, inparticular, active ingredients, having a relative concentration in therange of 0.3 to 0.5%.

As discussed above, the emission (fluorescence) radiation is detected bythe radiation transmission means 22 and at least a first signalindicative of the sample fluorescence characteristics is communicated tothe analyzer 28. According to the invention, the emission radiation isthen compared to the stored fluorescence characteristics of knownelements to identify the element or elements (or trace element(s)) inthe samples 14. The concentration of the element(s) can also bedetermined through the formulations referenced above (e.g., Eq. 3).

As also indicated above, the fluorescence detection means 20 is furtheradapted to be in synchrony with the conveyor system 50. In a preferredembodiment of the invention, the fluorescence detection means 20includes second control means 29 that is in communication with the firstcontrol means 24 and conveyor system 50. The second control means 29 isdesigned and adapted to synchronize the movement of the samples 14 onthe conveyor system 50 with the illumination of each sample 14 as ittraverses a respective sample path SP₁, SP₂. Thus, 100% inspection ofeach sample 14 contained in the blisters 12 is ensured.

Further, the noted synchronized sample fluorescence detection andanalysis is preferably accomplished at a rate (or speed) ofapproximately 1 sample/sec. Thus, the method and system of the inventionprovides high speed, accurate, on-line analysis of MDPI formulations andother pharmaceutical compositions that is unparalleled in the art.

The present invention will now be illustrated with reference to thefollowing examples. The examples are provided for illustrative purposesonly, and are not intended to limit the scope of the invention.

EXAMPLE 1

A MDPI formulation comprising >99.5% lactose and <0.5% active ingredientwas prepared. Referring to FIG. 9, the MDPI formulation and a referencelactose sample were then subjected to a pre-determined, suitable rangeof incident radiation to induce a fluorescent response. As will beappreciated by one having ordinary skill in the art, the incidentradiation is determined by and, hence, dependent upon the targetingredient or element of the MDPI formulation.

As illustrated in FIG. 9, a definitive fluorescent response, reflectingthe detection of the active ingredient was provided with an incidentradiation level in the range of approx. 350 mn to 500 nm. The notedfluorescence spectra further indicates that an active ingredient ortrace element having a relative concentration of less than 0.5% canreadily be detected by virtue of the fluorescence detection means of theinvention.

As will be appreciated by one having ordinary skill in the art, thenoted fluorescence spectra can be compared to stored calibration (orreference) spectra by conventional means to identify the detected activeingredient (or trace element). Further, as discussed above, theconcentration of the detected active ingredient can also be determinedthrough known formulations (See Eq. 3).

Applicants have further found that subjecting the MDPI formulation tosubsequent incident radiation in the same range provides little, if any,variation in the detected emission radiation. Indeed, the fluorescencespectra obtained were virtually identical.

Accordingly, by virtue of the fluorescence detection means of theinvention, a tolerance level of ±0.5 nm (i.e., calibration emissionradiation ±0.5 mn) can be employed. As will be appreciated by one havingordinary skill in the art, the noted tight “QC” specification isunparalleled in the art.

EXAMPLE 2

Referring now to FIG. 10, there are shown the fluorescence spectra ofsimilar MDPI formulations having ˜0.43% active ingredient (Curve A);˜0.42% active ingredient (Curve B); ˜0.41% active ingredient (Curve C);˜0.39% active ingredient (Curve D); and ˜0.37% active ingredient (CurveE). The noted fluorescence spectra were similarly induced with anincident radiation level in the range of approximately 350 to 500 nm.

The fluorescence spectra (i.e., Curves A–E) further demonstrate that asharp, definitive fluorescent response can be achieved in activeingredients having a relative concentration in the range of approx.0.37% to 0.43% by virtue of the fluorescence detection means of theinvention.

As will be appreciated by one having ordinary skill in the art, anarrower band or range of incident radiation (e.g., 375–475 nm) couldalso be employed to identify and determine the relative concentration ofan active ingredient. Further, an even narrower range of incidentradiation wavelengths (e.g., 400–425 nm) or incident radiation with asingle wavelength within the noted range (e.g., 410 nm) could beemployed to determine active ingredient “presence”.

SUMMARY

From the foregoing description, one of ordinary skill in the art caneasily ascertain that the present invention provides a method and systemfor high speed, real-time, 100% fluorescent inspection of MDPIformulations and other pharmaceutical compositions. The method andsystem of the present invention further provides an accuratedetermination of (i) the presence (i.e., qualitative assessment), and(ii) identity and concentration (i.e., quantitative assessment) ofactive ingredients and/or other trace elements having a relativeconcentration in the range of approximately 0.3 to 0.5%

Without departing from the spirit and scope of this invention, one ofordinary skill can make various changes and modifications to theinvention to adapt it to various usage and conditions. As such, thesechanges and modifications are properly, equitably, and intended to be,within the full range of equivalence of the following claims.

1. A system for use in in-situ analysis of pharmaceutical samples, saidsystem comprising: means for holding a plurality of said samples,wherein said samples are present in the form of a Metered Dry PowderInhaler formulation; means for moving said plurality of samples along asample path; means for generating a plurality of incident radiationpulses of different wavelength; means for illuminating at least arespective one of said samples with at least a respective one of saidradiation pulses during said movement of said samples, said radiationpulse having a suitable range of radiation wavelengths capable ofinducing a fluorescent response; means for detecting a first resultantfluorescence emitted from each of said samples; first control means incommunication with said moving means and said incident radiationgenerating means for synchronizing said means for illuminating each ofsaid samples with said moving means.
 2. The system of claim 1, includingsecond control means for analyzing second resultant fluorescence emittedfrom each of said samples.
 3. The system of claim 1, wherein saidplurality of incident radiation pulses of different wavelengths is inthe range of 200 to 800 nm.
 4. A system for use in determining thepresence and concentration of trace elements in a sample, said systemcomprising: means for holding a plurality of said samples, wherein saidsamples are present in the form of a Metered Dry Powder Inhalerformulation, each of said plurality of samples including at least one ofsaid trace elements; means for moving said plurality of samples along asample path; means for generating a plurality of incident radiationpulses of different wavelengths; means for illuminating at least arespective one of said samples with at least a respective one of saidradiation pulses during said movement of said samples, said radiationpulse having a suitable range of fluorescence radiation wavelengthscapable of inducing a fluorescent response; means for detecting aresultant fluorescence response emitted from said trace element; andfirst control means in communication with said moving means and saidincident radiation generating means for synchronizing said means forilluminating each of said samples with said moving means.
 5. The systemof claim 4, including second control means for storing fluorescencecharacteristics of pre-determined elements and means for comparing saiddetected resultant fluorescence emitted from said trace element toidentify said trace element in said plurality of samples, said secondcontrol means including means for determining the relative concentrationof said trace element in each of said samples.
 6. The system of claim 4,wherein said trace element has a relative concentration in the range 0.3to 0.5%.
 7. The system of claim 4, wherein said plurality of incidentradiation pulses of different wavelength range from 200 to 800 nm.
 8. Asystem for use in in-situ analysis of pharmaceutical compositionsamples, said system comprising: means for holding a plurality ofsamples, wherein said samples are present in the form of a Metered DryPowder Inhaler formulation, said samples including at least one traceelement; means for substantially simultaneously moving said plurality ofsamples along a sample path, illuminating at least a respective one ofsaid samples with incident radiation having one or more suitablewavelengths during said movement of said plurality of samples, anddetecting a result in emission radiation from said samples; and controlmeans in communication with said illuminating and detecting means forproviding a range of radiation and analyzing said result in emissionradiation and fluorescence emitted from said samples.
 9. The system ofclaim 8, wherein said incident radiation is directed along a firstradiation path that intersects said sample path and is substantiallyperpendicular thereto.
 10. The system of claim 9, wherein said emittedradiation is substantially detected along a second radiation path, saidsecond radiation path being substantially coincident with said firstradiation path.
 11. The system of claim 8, wherein said samples aremoved by said moving means at a minimum rate of one sample per second.12. The system of claim 8, wherein said incident radiation has aplurality of different wavelengths in the range of 200 to 800 nm. 13.The system of claim 8, wherein said trace element has a relativeconcentration in the range of 0.3 to 0.5%.